The p21/CDKN1/WAF1/CIP1 protein (El-Deiry, W. S.; et al. Cell 75: 817-825, 1993; Harper, J. W.; et al. Cell 75: 805-816, 1993; Huppi, K et al. Oncogene 9: 3017-3020, 1994) is a cell cycle control protein that inhibits cyclin-kinase activity, is tightly regulated at the transcriptional level by p53, and mediates p53 suppression of tumor cell growth. Along with p53, p21 appears to be essential for maintaining the G2 checkpoint in human cells (Bunz, F.; Dutriaux, A.; et al. Science 282:1497-1501, 1998). Sequences of P21 are well-conserved throughout evolution, and have been identified in species as diverse as human (Genbank Identifier 13643057), Drosophila melanogaster (GI#1684911), Caenorhabditis elegans (GI#4966283), and yeast (GI#2656016).
RORA (retinoic acid receptor related orphan receptor A) is a member of the nuclear hormone receptor superfamily (Giguere, V., et al (1994) Genes And Development 8: 538-53). Members of this gene family include the steroid hormone, thyroid hormone and retinoid receptors, and orphan receptors for which a ligand has not yet been identified. Members of this superfamily also share a common modular structure composed of a transactivation domain, a DNA-binding domain, and a ligand-binding domain. Typically, their transcriptional transactivation function is regulated by small lipophilic molecules, such as steroid hormones, vitamin D, retinoic acids, and thyroid hormone. These molecules are synthesized in the organism and pass readily through the plasma membrane to reach the corresponding receptors inside the cell.
Mutations in the RORA gene have been related to recessive Robinow syndrome (Afzal, A. R., et al (2000) Nat Genet 25:419-22). RORB is a transcription factor and interacts with NM23-2, a nucleoside diphosphate kinase involved in organogenesis and differentiation (Paravicini, G. et al. (1996) Biochem. Biophys. Res. Commun. 227: 82-87). RORC (ROR-gamma) is important for lymphoid organogenesis and plays an important regulatory role in thymopoiesis (Kurebayashi, S. et al (2000) Proc. Nat. Acad. Sci. 97: 10132-10137).
The ability to manipulate the genomes of model organisms such as Drosophila provides a powerful means to analyze biochemical processes that, due to significant evolutionary conservation, have direct relevance to more complex vertebrate organisms. Due to a high level of gene and pathway conservation, the strong similarity of cellular processes, and the functional conservation of genes between these model organisms and mammals, identification of the involvement of novel genes in particular pathways and their functions in such model organisms can directly contribute to the understanding of the correlative pathways and methods of modulating them in mammals (see, for example, Mechler B M et al., 1985 EMBO J 4:1551-1557; Gateff E. 1982 Adv. Cancer Res. 37: 33-74; Watson K L., et al., 1994 J Cell Sci. 18: 19-33; Miklos G L, and Rubin G M. 1996 Cell 86:521-529; Wassarman D A, et al., 1995 Curr Opin Gen Dev 5: 44-50; and Booth D R. 1999 Cancer Metastasis Rev. 18: 261-284). For example, a genetic screen can be carried out in an invertebrate model organism having underexpression (e.g. knockout) or overexpression of a gene (referred to as a “genetic entry point”) that yields a visible phenotype. Additional genes are mutated in a random or targeted manner. When a gene mutation changes the original phenotype caused by the mutation in the genetic entry point, the gene is identified as a “modifier” involved in the same or overlapping pathway as the genetic entry point. When the genetic entry point is an ortholog of a human gene implicated in a disease pathway, such as p21, modifier genes can be identified that may be attractive candidate targets for novel therapeutics.
All references cited herein, including patents, patent applications, publications, and sequence information in referenced Genbank identifier numbers, are incorporated herein in their entireties.